In preliminary studies, I found that treatment of Chinese hamster ovary cells with toxic doses of some alkylating agents makes the surviving population permanently more resistant to the toxic effects of these agents. Resistance does not seem to occur by gene mutation or selection, but rather by some other process. I propose to define the process by which increased resistance is achieved in more detail. A resistant clone which is representative of the resistant population and which has been stable in culture for 11 months will be characterized with respect to its mutabililty by alkylating agents and its repair capacity. The frequency at which resistant clones occur at different treatment doses will be determined by use of a replica plating method, and more clones will be tested in their response to alkylating agents to find out if they are equally resistant or if resistance increases with pretreatment dose. Also, more alkylating agents will be tested to establish which structural characteristics are necessary to increase resistance in progeny cells. Finally, it will be determined if increased resistance after treatment with alkylating agents occurs in other cell lines if it depends on their initial sensitivity to alkylating agents. The results will increase our understanding of how lakylating agents cause cell killing and what tolerance mechanisms are available to the mammalian cell to overcome the toxic effects of agents.